基于微孔过滤板的蛋白吸附高通量筛选方法

发布时间：2018-06-15 06:54

本文选题：高通量筛选 + 微孔过滤板　；参考：《化工学报》2017年06期

【摘要】：针对色谱分离过程优化,建立了基于微孔过滤板的蛋白吸附高通量筛选方法,用于介质初筛、吸附性能考察、吸附等温线和吸附动力学测定、吸附和洗脱条件优化等。首先优化了96孔过滤板的操作参数,以2种离子交换介质和2种混合模式介质为典型代表,采用微孔过滤板方法考察了不同介质和液相条件下牛血清白蛋白的吸附,得到结合载量分布图,确定了合适的蛋白吸附和解吸条件。进一步测定了4种介质在特定吸附条件下的吸附等温线和吸附动力学曲线,获得吸附相关参数。最后,采用微孔过滤板进行了洗脱条件优化,并与填充柱色谱分离进行比较,验证了方法的可靠性。结果表明,基于微孔过滤板的蛋白吸附高通量筛选是切实可行的,可以快速筛选色谱介质和液相,优化蛋白分离条件,具有资源消耗小、实验通量大、研发周期短、适用性广、稳定性高的特点,是蛋白色谱分离过程优化的一种新方法。
[Abstract]:According to the optimization of chromatographic separation process, a high throughput screening method for protein adsorption based on microporous filter plate was established, which was used for medium screening, adsorption performance evaluation, adsorption isotherm and adsorption kinetics determination, and optimization of adsorption and elution conditions. Firstly, the operating parameters of 96 hole filter plate were optimized. The adsorption of bovine serum albumin (BSA) in different media and liquid phase was investigated by using two kinds of ion exchange media and two mixed mode media as typical representatives. The suitable conditions of protein adsorption and desorption were determined. The adsorption isotherms and adsorption kinetics curves of four kinds of media under specific adsorption conditions were further determined and the adsorption parameters were obtained. Finally, the elution conditions were optimized by using microporous filter plate and compared with packed column chromatography to verify the reliability of the method. The results show that the high throughput screening of protein adsorption based on microporous filter plate is feasible. It can quickly screen chromatographic medium and liquid phase, optimize the conditions of protein separation, have the advantages of low resource consumption, large experimental flux, short R & D cycle and wide applicability. Because of its high stability, it is a new method to optimize the separation process of protein chromatography.
【作者单位】：浙江大学化学工程与生物工程学院生物质化工教育部重点实验室;
【基金】：国家自然科学基金项目(21476198,21576233) 国际科技合作专项项目(2015DFG42070)~~
【分类号】：O652.6;TQ936.2

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